Cannabidiol regulaton of learned fear: role of endocannabinoids

Anxiety-related disorders like post-traumatic stress disorder (PTSD) affect 1 in 4 people in their lifetime. PTSD is characterized by abnormal fear memory persistence and inadequate treatment options given the temporary effectiveness of psychological therapies and side effects of medicines. Cannabidiol, the main non-psychotropic phytocannabinoid in cannabis, is safe for use and has both anxiolytic and cognitive enhancing properties. We have shown in rats that cannabidiol reduces learned fear but also enhances its extinction (Song et al., 2016), the psychological process underlying fear reduction in exposure therapy. These properties make cannabidiol an attractive candidate drug for treating PTSD but its underlying molecular mechanisms remain unclear. Cannabidiol regulation of extinction is blocked by cannabinoid receptor type1 antagonist pretreatment. This suggests that its effects are mediated indirectly by elevating brain endocannabinoid levels, although which endocannabinoids are involved remains unknown. Moreover, cannabidiol may act in combination with other phytocannabinoids in cannabis to have synergistic effects on learned fear and extinction, possibly also via endocannabinoid signaling.

Aim, Objectives and Hypothesis:
We will determine the role of endocannabinoids in mediating the effects of cannabidiol, and a novel proprietary phytocannabinoid drug combination, on learned fear and extinction in rats.
Objective 1: We will quantify ex vivo levels of endocannabinoids (e.g. 2-arachidonoylglycerol, anandamide) in brain regions of interest after cannabidiol treatment followed by extinction.
Objective 2: We will determine the effects of a novel proprietary phytocannabinoid drug combination on (1) learned fear and extinction, and (2) ex vivo levels of brain endocannabinoids after drug treatment followed by extinction.

Hypothesis: Enhanced extinction caused by cannabidiol, and potentially also a novel phytocannabinoid drug combination, will result in elevated brain endocannabinoid levels.

Methods:
Drug administration and behavioural testing: Rats will undergo fear conditioning and extinction, with cannabidiol or a novel phytocannabinoid drug combination (or vehicle) injected before extinction; some rats will also undergo later extinction testing drug-free. 'Freezing' behaviour will be quantified as a measure of learned fear during extinction and extinction testing.

Endocannabinoid quantification: Some rats will be culled after extinction and their brains will be removed to dissect regions of interest for later endocannabinoid quantification using liquid chromatography-tandem mass spectrometry (LC-MS).

Innovation:
Although the pharmacological mechanisms underlying the effects of cannabidiol on fear regulation have begun to be elucidated, the underpinning molecular mechanisms remain poorly understood. We will use cutting-edge LC-MS techniques to profile the levels of endocannabinoids in the brain. We will also compare the effects of cannabidiol and a novel phytocannabinoid drug combination to determine if the latter has a greater effect than cannabidiol alone. This will potentially lead to the use of phytocannabinoids to treat anxiety in the future.